3D bioprinting of dual-crosslinked nanocellulose hydrogels for tissue engineering applications

Hydrogels based on cellulose nanofibrils (CNFs) have been widely used as scaffolds for biomedical applications, however, the poor mechanical properties of CNF hydrogels limit their use as ink for 3D bioprinting in order to generate scaffolds for tissue engineering applications. In this study, a dual crosslinkable hydrogel ink composed of a poly(ethylene glycol) (PEG) star polymer and 2,2,6,6-tetramethyl-1-piperidinyloxy (TEMPO)-oxidized nanocellulose fibers (CNFs) is presented. As the resulting hydrogel had low structural integrity, at first crosslinking of CNFs was introduced by Ca²⁺. Strong physical interactions between CNFs and Ca²⁺ cations allowed easy regulation of the viscosity of the inks for extrusion printing raising the solution viscosity by more than 1.5 times depending on the amount of Ca²⁺ added. The resulting hydrogel had high structural integrity and was further stabilized in a second step by photo crosslinking of PEG under visible light. In only a few seconds, hydrogels with Young's modulus between ∼10 and 30 kPa were obtained just by altering the CNF and Ca²⁺ content. 3D printed hydrogels supported fibroblasts with excellent cell viability and proliferation. The dual crosslinkable hydrogel ink herein developed is versatile, easy to prepare, and suitable for 3D printing of bioscaffolds with highly tailored viscoelastic and mechanical properties applicable in a wide range of regenerative medicines.